2163. Prevalence and geographic distribution of mutations in TonB-dependent receptors in the Prospective Observational Pseudomonas aeruginosa (PA) Study

Abstract Background Cefiderocol (FDC) is a last line treatment for carbapenem-resistant Pseudomonas aeruginosa (CR-PA). FDC retains activity against CR-PA by inhibition of penicillin-binding proteins facilitated by active uptake of drug via TonB-dependent receptors (TBDR). Prior, we have shown an association between mutations in TBDR genes and loss of FDC susceptibility, particularly in association with changes at the PA-derived cephalosporinase (PDC/AmpC) active site. Here, we outline geographic variation and prevalence of TBDR mutations in the Prospective Observational Pseudomonas (POP) study.Figure 1. SNP frequency in the pirR, pirS, pirA, and piuA/piuD genes across geographic location.Figure 2. Genotypic variation of PirRS and PiuA/D genes in the 972 P. aeruginosa isolates of the POP cohort. Black versus gray differentiates presence of either PiuA versus PiuD homologous proteins. Color legend corresponds to presence of TBDR gene variants/SNPs per gene pirR, pirS, pirA, piuA/piuD, and ampC. Methods The genomes of 972 CR-PA isolates collected across 10 countries in North America, South America, Asia, and Oceania were screened for single nucleotide polymorphisms (SNP) resulting in missense, insertion/deletion (indel), frameshift, and early stop codon mutations in the TBDR genes pirR, pirS, pirA, piuA/piuD, and ampC using PAO1 as a reference. SNPs were defined as variant mutations occurring in ≤10% of isolates. A core genome maximum likelihood phylogenetic tree was used to assess distribution and prevalence of genotypic variants. Neighbor-joining trees of the variants in each gene were used to analyze clusters across lineages. Results Across all regions except Oceania, SNPs were most frequently seen in pirS and pirA (Fig 1). Gene variants comprised from 8% up to 50% of each geographic population. On phylogenetic mapping, isolates within the same lineage tended to have clustering of shared mutations across the TBDR genes pirS, pirR, and pirA, and the majority of piuA/piuD mutations clustered within two distinct lineages (Fig 2). Major mutations, including indel, frameshift, and early stop codon mutations occurred in 16 isolates (1.6%), predominantly in isolates from the United States (n=14). Variations in PDC were frequently observed, with 57 different variants across 35.8% of the population. Seven of the PDC variants (n=10 isolates) have been associated with decreased susceptibility to ceftolozane/tazobactam, but these variants did not co-occur with major TBDR mutations. Conclusion TBDR mutations occur in PA isolates worldwide. Mutations previously associated with reduced susceptibility to FDC occurred most frequently in isolates from the United States. Disclosures Michael J. Satlin, MD, AbbVie: IDMC member|Biomerieux: Grant/Research Support|Merck: Grant/Research Support|Shionogi: Advisor/Consultant|SNIPRBiome: Grant/Research Support William R. Miller, M.D., Merck: Grant/Research Support|UpToDate: Honoraria

Background.Cefiderocol (FDC) is a last line treatment for carbapenem-resistant Pseudomonas aeruginosa (CR-PA).FDC retains activity against CR-PA by inhibition of penicillin-binding proteins facilitated by active uptake of drug via TonB-dependent receptors (TBDR).Prior, we have shown an association between mutations in TBDR genes and loss of FDC susceptibility, particularly in association with changes at the PA-derived cephalosporinase (PDC/AmpC) active site.Here, we outline geographic variation and prevalence of TBDR mutations in the Prospective Observational Pseudomonas (POP) study.Methods.The genomes of 972 CR-PA isolates collected across 10 countries in North America, South America, Asia, and Oceania were screened for single nucleotide polymorphisms (SNP) resulting in missense, insertion/deletion (indel), frameshift, and early stop codon mutations in the TBDR genes pirR, pirS, pirA, piuA/piuD, and ampC using PAO1 as a reference.SNPs were defined as variant mutations occurring in ≤10% of isolates.A core genome maximum likelihood phylogenetic tree was used to assess distribution and prevalence of genotypic variants.Neighbor-joining trees of the variants in each gene were used to analyze clusters across lineages.
Results.Across all regions except Oceania, SNPs were most frequently seen in pirS and pirA (Fig 1).Gene variants comprised from 8% up to 50% of each geographic population.On phylogenetic mapping, isolates within the same lineage tended to have clustering of shared mutations across the TBDR genes pirS, pirR, and pirA, and the majority of piuA/piuD mutations clustered within two distinct lineages (Fig 2).Major mutations, including indel, frameshift, and early stop codon mutations occurred in 16 isolates (1.6%), predominantly in isolates from the United States (n=14).Variations in PDC were frequently observed, with 57 different variants across 35.8% of the population.Seven of the PDC variants (n=10 isolates) have been associated with decreased susceptibility to ceftolozane/tazobactam, but these variants did not co-occur with major TBDR mutations.
Conclusion.TBDR mutations occur in PA isolates worldwide.Mutations previously associated with reduced susceptibility to FDC occurred most frequently in isolates from the United States.
Disclosures.Michael J. Satlin, MD, AbbVie: IDMC member|Biomerieux: Grant/Research Support|Merck: Grant/Research Support|Shionogi: Advisor/ Consultant|SNIPRBiome: Grant/Research Support William R. Miller, M.D., Merck: Grant/Research Support|UpToDate: Honoraria Resistant population was calculated by dividing the cfu/mL on C/T containing plates by the cfu/mL on drug free agar (total population).Percentage of C/T-resistant population was compared between MEM-exposed and MEM non-exposed passages of each isolate.

Figure 2 .
Figure 2. Genotypic variation of PirRS and PiuA/D genes in the 972 P. aeruginosa isolates of the POP cohort.Black versus gray differentiates presence of either PiuA versus PiuD homologous proteins.Color legend corresponds to presence of TBDR gene variants/SNPs per gene pirR, pirS, pirA, piuA/piuD, and ampC.

1
University of Houston College of Pharmacy, Houston, Texas 2 Department of Pharmacy Practice and Translational Research, University of Houston College of Pharmacy, Houston, Texas, USA, Houston, TX 3 The University of Texas Health Science Center, Houston, Texas 4 Rice University, Houston, Texas 5 Broad Institute of MIT and Harvard, Cambridge, Massachusetts 6 University of Houston, Houston, TX 1 Hartford Hospital, Farmington, Connecticut 2 Sanofi, Boston, Massachusetts 3 Johns Hopkins School of Medicine, Baltimore, MD 10 cfu/mL).Samples were plated on drug free and drug containing agar (C/T concentration 16 mg/L) where growth on C/T agar indicated resistance development.